Abstract
Purpose :
To analyze the effects of long-pulsed, 1 MHz ultrasonic application on human retinal pigment epithelium (ARPE-19) cell viability at various doxorubicin concentrations.
Methods :
In this in vitro study, ARPE-19 cells were plated in 96-well plates and treated with 360 μL of solution containing 0.01, 0.1, 1, 10, or 100 μg/mL of doxorubicin. 5.5 mg of doxorubicin (Fisher Scientific) was first dissolved in 550 μL of DMSO to form a stock solution. The stock solution was then transferred to 54.45 mL of media and serially diluted to create the various titrated concentrations. Cell plates were sealed with MicroAmpTM Optical Adhesive Film, inverted, and placed in a water bath at room temperature. Cell plates were then treated with 1 MHz of ultrasound (US) at a power of 4.93 W/cm2 and pulse repetition frequency (PRF) of 30 Hz for five minutes (Olympus EPOCH 650 Ultrasound). Media were removed from the wells and replaced with 100 μL of media with the corresponding doxorubicin concentration. The control mimicked all parameters including the 1% DMSO but cells did not receive US application. As an additional control group, US was applied to cells without doxorubicin to analyze the effects of US on cell viability. MTT assays were performed at both 24 and 48 hours to quantify cellular metabolism, which had been used in literature as a marker for cell viability.
Results :
At 24 hours after US was applied, there was a significant decrease in viability in cells treated with doxorubicin 0.1 µg/mL to 100 µg/mL (Figure 1). The same trend was observed at 48 hours (Figure 2). At both 24 and 48 hours, the untreated group without doxorubicin showed that ultrasound alone did not have a significant effect on cell viability (rightmost two columns in Figures 1 and 2).
Conclusions :
US has played a pivotal diagnostic role in ophthalmology, but its therapeutic potential has been largely unexplored. This study demonstrated that long-pulsed US enhanced the cytotoxic effect of doxorubicin at physiologically relevant concentrations. This observation suggests a potential adjunctive role of ultrasound in treating tumors. Further studies using other cell types are underway to further explore the therapeutic role of ultrasound.
This is a 2021 ARVO Annual Meeting abstract.